Analysis Of CircRNA Expression Profile In Trisomy 13 Syndrome And Construction Of CircRNA-related CeRNA Network

13-Trisomatic syndrome,also known as Patau syndrome?PS?,is a disease caused by the patient's egg cells do not separate on chromosome 13 during the first meiosis.The clinical symptoms and multiple malformations of patients with PS are more serious than those of trisomy 18 and trisomy 21.They often show abnormalities of nervous system,immune system,urogenital system and congenital cardiac hypoplasia,accompanied by severe mental retardation,bradykinesia,dyslexia and polydactyly.The birth of patients with PS has brought enormous mental pressure and heavy financial burden to family and society.At present,there is no effective medical method for PS patients,so pregnant women need effective prenatal screening and prenatal diagnosis for birth intervention to avoid the birth of children with PS.However,traditional prenatal screening is harmful to maternal and infant health due to the invasive sampling method,and the existing noninvasive prenatal diagnosis methods have such problems as high cost and cumbersome operation.Therefore,it is of great clinical significance to explore new noninvasive prenatal screening markers for trisomy 13 syndrome.Objective: To sequence the whole transcriptome of pregnant women with trisomy 13 syndrome and normal pregnant women by using RNA-seq technology from the perspective of circRNA,so as to lay a foundation for exploring differential expression circRNA as a prenatal screening marker for trisomy 13 syndrome and its related ceRNA regulatory network.Methods: Umbilical cord blood and peripheral blood of patients diagnosed as 13-trisomy syndrome by prenatal diagnosis were extracted as the experimental group,and umbilical cord blood and peripheral blood of two pregnant women with normal pregnancy and the same gestational age were extracted as the control group.Total RNA was extracted by Trizol method.RNA integrity and RNA concentration were detected by NanoDrop and Agilent 2100,respectively.The expression profiles of circRNA and RNA in samples were obtained by RNA-seq technique and bioinformatics analysis was carried out.The binding sites of circRNA and microRNA were predicted based on the software of Mianda.The differential expression circRNA and RNA in umbilical cord blood on chromosome 13 were verified by real-time fluorescence quantitative PCR.Finally,a ceRNA network of umbilical cord blood with 13-trisomy syndrome was constructed by the combined analysis of circRNA,microRNAs and RNA with differential expression in umbilical cord blood,in order to explore the biological function of circRNA in trisomy 13 syndrome and its regulation mechanism.The results were as following:?1?13437 circRNAs were detected in umbilical cord blood samples of the two groups,of which 11781 circRNAs showed statistical significance,including 4458 up-regulated circRNAs and 7323 down-regulated circRNAs.A total of 27 circRNA expressed differentially on chromosome 13 of umbilical blood and peripheral blood,including 15 circRNAs significantly upregulated,the most up-regulated circRNA was hg38_circ₀005726,of which the expression difference multiple was 6.69;12 circRNAs significantly down-regulated,the most down-regulated circRNA was hsa_circ₀030567,of which the expression difference multiple was-7.12.The expression of circRNA on chromosome 13 in umbilical cord blood was verified by qRT-PCR in peripheral blood.The results showed that the differential expressions of hg38_circ₀005566,hsa_circ₀000502,hsa_circ₀005651 and hsa_circ₀030567 were statistically significant.?2?A total of 19715 differential expression genes were detected in umbilical cord blood samples of the two groups,of which 14003 differential expression genes have biological and statistical significance.EEF1A1 was the most differentially expressed gene in 8728 up-regulated genes,and CTSB was the most differentially expressed gene in 5275 down-regulated genes.The expression of SKA3,VWA8,ITM2 B and ABCC4 genes in peripheral blood was verified by qRTPCR.The results showed that the differential expression of SKA3,VWA8 and ITM2 B had statistical significance?P < 0.05?,but the differential expression of ABCC4 had no statistical significance?P > 0.05?.?3?By constructing ceRNA network differentially expressed by cord blood,it was found that hg38_circ₀005566,hsa_circ₀000502,hsa_circ₀005651 may regulate the expression of SKA3,VWA8 and ITM2 B through miRNA sponge;in addition,hsa_circ₀000502,hg38_circ₀005666,hsa_circ₀0030567 can jointly regulate the expression of miR-454-3p,possibly in PS.Conclusion:?1?There are a large number of differentially expressed circRNAs in cord blood and peripheral blood of PS patients.?2?The binding of circRNA to miRNA plays a sponge role and may play an important role in the regulation of PS gene expression.?3?Bioinformatics analysis of differential expression genes showed that these genes were mainly involved in the process of nucleic acid binding and protein binding,as well as T cell receptor signaling pathway and endometrial cancer,etc.This may be related to fetal immune system,nervous system and cardiovascular system development defects in patients with PS.?4?TThe differentially expressed hg38_circ₀005566,hsa_circ₀000502,hsa_circ₀005651 and their target genes SKA3,VWA8,and ITM2 B may be new biomarkers for non-invasive prenatal screening diagnosis of PS.